The Function Of Promoter Hypermethylation Of The P15,P16,HIC-1 Genes In The Occurrence And The Malignant Transformation Process Of Hydatidiform Mole

Objective: Gestational trophoblastic disease (GTD) is a group of diseases from placental trophoblast cells, including hydatidiform mole, invasive mole, choriocarcinoma and placental site trophoblastic tumor. The reported incidence of GTD is about two to ten times higher in Asia than that in United States and Europe, which seriously impact on women's reproduction and health. Hydatidiform mole is one major subtype of GTD, represents abnormally formed placenta characterized by swelling of chorionic villi and excessive proliferation of trophoblast. Although not a frank malignancy, hydatidiform mole may behave in an aggressive way with a predisposition for malignant transformation, and lead to invasive mole or choriocarcinoma. Nowadays, the occurred and malignant transformation mechanisms of hydatidiform mole are not clear. The mainstays for predicting the development of hydatidiform mole largely dependent on serial serum human chorionic gonadotrophin (HCG) titers and the clinical symptom and physical sign, except that there are no better objective indexes for predicting the malignant transformation of hydatidiform mole. The p15 and p16 gene encode proteins belonging to the cyclin-dependent-kinase-inhibit family, inhibit Cdk4/6 and Cyclin D association, thus prevent the cell entering the S phase from the G1 phase, and show the tumor suppressor genes'function. The two genes play an important role in a variety of human tumors, and their inactivation mechanisms are loss,mutation and methylation of the promoter region. The hypermethylated in cancer-1 (HIC-1), a new candidate tumor suppressor gene located in 17p13.3, encodes a protein with five Krüppel-like C2H2 zinc finger motifs and a N-terminal protein-protein interaction domain called BTB/POZ. The wild-type p53 gene has been widely implicated in the regulation of HIC-1 transcription. AS a master growth regulatory gene, the absence or inactivation of HIC-1 gene is closely related to a variety of human cancers. At present, the roles of hypermethylation of the p15,p16 and HIC-1 genes in hydatidiform mole have reported fewly, so researches on the promoter hypermethylation of the three genes will contribute to a futher understanding of the molecular mechanisms in occurred and malignant transformation of hydatidiform mole. In this study, we examined the frequency of methylation of p15,p16 and HIC-1 genes in 30 hydatidiform moles and 20 normal first trimester placentas by methylation-specific polymerase chain reaction (MSP), in order to discuss the roles of the promoter hypermethylation of p15,p16 and HIC-1 genes in the occurrence and the malignant transformation process of hydatidiform mole.Methods: The 30 specimens of hydatidiform mole were collected from the patients with hydatidiform mole after being suction evacuated the first time, at the Department of Obstetrics and Gynecology in Number Three Hospital of Hehei Medical University from January 2005 to May 2008, and were fixed in 10% formalin, including 19 cases of spontaneous regressive mole and 11 cases developed gestational trophoblastic neoplasia requiring chemotherapy. All the specimens were diagnosised by experienced pathologists, with a complete clinical and pathological data. The 20 normal first trimester placentas were obtained from patients need induced abortion because of pregnancy, as normal control, at out-patient clinic at the same time,and were fixed in 95% alcohol. Genomic DNA of the 30 hydatidiform moles and 20 normal first trimester placentas were extracted by phenol-chloroform-isoamyl alcohol, and the promoter methylation state of the p15, p16 and HIC-1 gene in these samples were detected by MSP. Statistical analysis was performed using the Statistical Package for Social Science 13.0 for Windows. Differences between groups were analyzed by the corrected chi-square or fisher's exact test where applicable. P values less than 0.05 was considered statistically significant. All statistical tests were calculated in two-sided.Results: 1. The methylation of p16 gene was not detected in normal placenta, whereas the frequency of the aberrant methylation of p16 gene in hydatidiform mole was 26.67%. There are significant deviation between them (P<0.05). Those hydatidiform moles, which subsequently developed gestational trophoblastic neoplasia, were also found to have significantly more frequent p16 promoter hypermethylation comparing to those that subsequently regressed (54.55% vs 10.53%,P<0.05). 2. Aberrant methylation of HIC-1 gene were detected in both normal placenta and hydatidiform mole, the frequency of the methylation of HIC-1 gene were 85.00% and 90.00%, respectively. But the difference between them was not statistically significant (P>0.05). The frequency of the methylation of HIC-1 gene in the malignant moles (81.82%) was lower than it in the spontaneous regressive mole (94.74%), but the difference was not statistically significant, either (P>0.05). 3. The methylation of p15 gene was not detected in normal placenta, the frequency of the methylation of p15 gene in hydatidiform mole was only 6.67%. There are no significant deviation between them (P>0.05).Conclusion: Promoter hypermethylation of the p16 gene might play an important role in the occurrence and the malignant transformation process of hydatidiform mole. Promoter hypermethylation of the HIC-1 gene might play a role in the early occurrence process,but not play clear role in the malignant transformation process. Promoter hypermethylation of the p15 gene might not play a clear role in neither the occurrence nor the malignant transformation processes.